Medicine, pharmacology and biology cannot, for various reasons, totally efrain from animalexperiments. This includes the necessity of painless anaesthesia and euthanasia of experimentalanimals. CO2 anaesthesia of rodents is a commonly used method, nevertheless, little is knownabout whether this method is painful to the animal. The present work should contribute toanswering the question of whether C O 2 anaesthesia of laboratory rats is compatible with thehumane treatment of laboratory animals.A method has been developed that can evaluate CO2 anaesthesia for symptoms of distress. As afirst step to this goal, suitable criteria for the assessment of distress were defined on the basis ofthe existing literature. The following experimental studies focused on two major aspects. First, thebehaviour of the animals during CO2 anaesthesia was monitored and checked for attemps toescape, retreat or attack as overt signs of distress. Secondly, blood ACTH, corticosterone andglucose levels were measured during anaesthesia as objective criteria of distress.In the first experiment 59 rats were anaesthesised with 2 L/min, 4 L/min and 6 L/min CO2,respectively. Three consecutive stages during anaesthesia were observed. An initial increasedrespiration rate was followed by immobility that eventually led to total muscle relaxation.The onset of these symptoms showed the smallest variability at 6 L/min CO2, so this flow rate waschosen for subsequent experiments.In order to measure objective distress parameters blood was taken during the course of CO2anaestesia. According to the observed stages samples were collected at 30 sec, 75 sec and 120sec, respectively. The CO2 concentrations at these timepoints were about 18%, 39% and 55%,respectively.48 animals were subdivided into four groups. Prior to CO2 anaesthesia rats either received anacepromazin sedative or were put under a general anaesthetic (pentobarbiturate). Control groupswere givenplacebos in the same way as the drugs used.The pretreated animals were compared to the controls during subsequent C02 anaesthesia. If C02would cause distress reactions, one would expect the unconscious animals to show less severealterations of the measured stress parameters. Contrary to this, neither a difference in behaviournor statistically significant alterations of the blood stress parameters could be found. Hence, theperformed experiments could not detect any symptoms of suffering during the C02 anaesthesia.Summarising, anaesthesia and euthanasia of laboratory rats using C02 can be evaluated, on thebasis of the obtained data, to be compatible with the humane treatment of animals. The methoddeveloped can be easily adapted to studies with other gas-based anaesthetics.